Comparison of Hepatic Metabolite Profiles between Infant and Adult Male Mice Using 1H-NMR-Based Untargeted Metabolomics

Abstract

Although age-related characteristics of hepatic metabolism are reported, those in infants are not fully understood. In the present study, we performed untargeted metabolomic profiling of the livers of infant (3-week-old) and adult (9-week-old) male ICR mice using ¹H-NMR spectroscopy and compared 35 abundant hepatic metabolite concentrations between the two groups. The liver/body weight ratio did not differ between the two groups; however, serum glucose, blood urea nitrogen, total cholesterol, and triglyceride concentrations were lower in infants than in adults. Hepatic carbohydrate metabolites (glucose, maltose, and mannose) were higher, whereas amino acids (glutamine, leucine, methionine, phenylalanine, tyrosine, and valine) were lower in infant mice than in adult mice. The concentrations of ascorbate, betaine, sarcosine, and ethanolamine were higher, whereas those of taurine, inosine, and O-phosphocholine were lower in infant mice than in adult mice. The differences in liver metabolites between the two groups could be due to differences in their developmental stages and dietary sources (breast milk for infants and laboratory chow for adults). The above results provide insights into the hepatic metabolism in infants; however, the exact implications of the findings require further investigation.

Keywords: 1H-NMR; adult; age difference; infant; liver; metabolite; metabolomics.

Figure 1

Body and liver weights and the ratio of liver/body weight of infant (3-week-old, n = 9) and adult (9-week-old, n = 9) male mice: (A) Body weight; (B) Liver weight; (C) Liver/body weight ratio. Data are presented as mean ± SE. Student’s t-test, *** p < 0.001, vs. infant.

Figure 2

Serum biomarkers of infant (3-week-old, n = 9) and adult (9-week-old, n = 9) male mice. Data are presented as mean ± SE. Student’s t-test, *,** p < 0.05, and 0.01, respectively, vs infant. ALT, alanine transaminase; AST, aspartate transaminase; ALB, albumin; Crea, creatinine; BUN, blood urea nitrogen; GLU, glucose; TC, total cholesterol; TG, triglyceride.

Figure 3

Representative 600 MHz 1H NMR spectra of the liver. Key: 1, isoleucine; 2, leucine; 3, valine; 4, lactate; 5, alanine; 6, Lysine; 7, ornithine; 8, acetate; 9, methionine; 10, glutamate; 11, glutamine; 12, carnitine; 13, glutathione; 14, ß-alanine; 15, dimethylamine; 16, sarcosine; 17, carnosine; 18, creatine; 19, ethanolamine; 20, choline; 21, O-phosphocholine; 22, sn-glycerol-3-phosphocholine; 23, taurine; 24, betaine; 25, glucose; 26, glycine; 27, maltose; 28, mannose; 29, ascorbate; 30; uridine; 31, inosine; 32, fumarate; 33, tyrosine; 34, phenylalanine; 35, niacinamide.

Figure 4

Boxplot of the statistically important hepatic metabolites (p < 0.05) in infant (3-week-old, n = 9) and adult (9-week-old, n = 9) male mice. Metabolites written in red letters are higher in the infants, while those in blue letters are higher in the adults.

Figure 5

PLS−DA and VIP score plots. (A) PLS−DA score plot comparing infant (3−week−old, n = 9) and adult (9−week−old, n = 9) male mice groups. (B) The variable importance in projection (VIP) score plot from PLS−DA analysis of important metabolites from infant and adult groups.

Figure 6

Heatmap hierarchical clustering analysis for top 20 metabolites that differed among infant (3−week−old, n = 9) and adult (9−week−old, n = 9) male mice. Red and blue colors indicate fold higher and lower relative concentrations of metabolites, respectively.

Figure 7

Metabolite sets enrichment analysis. The impacts and significances of biochemical pathways affected by different metabolite profiles of infant (3−week−old, n = 9) and adult (9−week−old, n = 9) male mice livers. The color and size of each node indicate the p value and pathway impact value, respectively. Darker red color indicates higher p value.